The tumor suppressor gene p53 is mutated in more than 50% of human tumors. The p53 protein has been described as the guardian of the genome because it responds to DNA damage or other types of stress by either arresting the cell in the G1 phase for damage repair or initiating an apoptotic pathway to eliminate the damaged cell entirely. However, the mechanisms that govern the transcriptional program of p53 are not well understood. In our early studies, we found that CBP/p300 promotes p53-dependent transcription by directly acetylating lysine residues in the C-terminal region of p53 (Lys 370, 371, 372, 381, 382 and 386). This study led to the notion that acetylation is a general protein modification for the regulation of non-histone proteins. Acetylation of p53 is also reversible in the presence of deacetylases such as HDAC1 and SIRT1 and the dynamic regulation of non-histone proteins by acetylation and deacetylation is now verified in the studies of many cellular factors. Tip60, a member of the MYST family of histone acetyltransferases was recently implicated in p53 activation through a large-scale RNAi screen. To elucidate the role of Tip60 in p53 regulation, we found that Tip60 directly interacts with p53 and also induces p53 acetylation specifically, at lysine 120 within the DNA binding domain. Interestingly, this modification is crucial for p53-dependent apoptosis but is dispensable for its mediated growth arrest. Lysine 120 is a recurrent site for p53 mutation in human cancer and the corresponding acetylation-defective tumor mutant (K120R) abrogates p53-mediated apoptosis but not growth arrest. Moreover, we have recently identified another acetylation site (K164) within the core domain. K164 acetylation is induced by CBP/p300 and this site is also mutated in human cancer. These studies implicate previous uncovered regulatory mechanisms of p53 and also suggest that the DNA binding domain, where the most tumor mutations reside, is critically involved in modulating p53 function. Further analysis of these modifications is required to clarify the precise role of p53 acetylation in transcription activation and yield broader insights into the mechanisms of p53 modifications in regulating its tumor suppressor function. In Aim 1, we will investigate the role of acetylation/deacetylation of the core domain in p53 regulation. In Aim2, we will identify and characterize novel factors in regulating K120 acetylation. Finally, in Aim 3, we will investigate the physiological role of K120 and other acetylation sites in p53 regulation. Overall, this study will reveal the precise roles of p53 acetylation/deacetylation in tumorigenesis. PUBLIC HEALTH RELEVANCE: The tumor suppressor gene p53 is mutated in more than 50% of human cancers. Post- translational modifications of the p53 protein are crucial for modulating p53 function in both normal cells and cancer cells. This study will characterize the roles of p53 acetylation/deacetylation in tumorigenesis and elucidate the precise mechanisms for Tip60-mediated acetylation in regulating the p53 pathway.